Location on the chromosome of Escherichia coli of genes governing purine metabolism

Jochimsen, Bjarne; Nygaard, Per; Vestergaard, Trygvi

doi:10.1007/bf00269424

Cited by 119 publications

(90 citation statements)

References 24 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…This strain lacks both the bacterial xanthine-guanine and hypoxanthine phosphoribosyltransferase activities (11). HGPRT expression was induced in low phosphate induction (LPI) medium as reported (10).…”

Section: Methodsmentioning

confidence: 83%

“…HGPRT Expression in E. coli and Purification of Recombinant Protein-L. donovani HGPRT was ligated into the pBAce expression vector and transformed into S⌽606 (⌬pro-gpt-lac, thi, hpt) E. coli as described (10,11). This strain lacks both the bacterial xanthine-guanine and hypoxanthine phosphoribosyltransferase activities (11).…”

Section: Methodsmentioning

confidence: 99%

“…Complementation Analysis-Wild type and mutant HGPRT-pSelect constructs were transformed into S⌽609 (⌬pro-gpt-lac, hpt, thi, pup, purHJ) E. coli, a purine auxotroph that lacks the same PRT activities as the SN606 cells (11). Transformants were isolated on LB plates containing 100 g/ml ampicillin and 50 g/ml streptomycin.…”

Section: Methodsmentioning

confidence: 99%

“…If either hypoxanthine or guanine is omitted from the medium, S⌽609 cells cannot survive because of the genetic defect (purHJ) in the purine biosynthetic pathway that confers purine auxotrophy. The adenine present in this minimal medium (see "Experimental Procedures") cannot serve as a source of guanylate nucleotides in the presence of high histidine levels in the media (11). In contrast, the S⌽609 cells transformants containing mutant hgprt genes exhibited three distinct growth phenotypes (Fig.…”

Section: Effects Of Protein Modifyingmentioning

confidence: 99%

See 3 more Smart Citations

The Conserved Serine-Tyrosine Dipeptide in Leishmania donovani Hypoxanthine-guanine Phosphoribosyltransferase Is Essential for Catalytic Activity

Jardim

Ullman

1997

Journal of Biological Chemistry

View full text Add to dashboard Cite

Protozoan parasites constitute a devastating public health and socioeconomic burden in the tropical and subtropical regions of the world. In the absence of effective vaccines, empirically derived drugs are the only means available to treat parasitic infections. However, the current arsenal of antiparasitic drugs is far from ideal, as these agents are moderately to highly toxic, possibly mutagenic and/or carcinogenic, and require protracted treatment regimens. The control of parasitic diseases has also been complicated by the emergence of drugresistant strains (1), further underscoring the need for novel antiparasitic agents. Rational development of new drugs that selectively target protozoan pathogens requires the exploitation of biochemical or metabolic differences between parasite and host. As protozoan parasites, unlike mammalian cells, are auxotrophic for purines and consequently rely on purine appropriation from the host for survival and proliferation (2), the purine salvage pathway has stimulated considerable interest as a paradigm for antiparasitic drug development. One enzyme that is central to purine acquisition in many parasites (2) is hypoxanthine-guanine phosphoribosyltransferase (HGPRT) 1 which catalyzes the phosphoribosylpyrophosphate (PRPP)-dependent phosphoribosylation of hypoxanthine and guanine to the nucleotide level. In some parasites, HGPRT also recognizes xanthine as a substrate (3) and is, therefore, termed a hypoxanthine-guanine-xanthine phosphoribosyltransferase (HGX-PRT). Genes and cDNAs encoding HG(X)PRT (HG(X)PRT) from a number of protozoan parasites have now been cloned, sequenced, and overexpressed in Escherichia coli, and the recombinant HG(X)PRT enzymes have been purified and characterized kinetically (3). Multisequence alignments demonstrate several short regions of amino acid homology among HG(X)-PRT family members that are flanked by much longer regions without significant sequence similarity. The crystal structures for the product-bound form of the human HGPRT (4) and the Tritrichomonas foetus HGXPRT (5) have revealed, however, a common core framework within the tertiary structure that is conserved among other members of the phosphoribosyltransferase (PRT) family (6 -8). These structures have established functional roles for all of the conserved regions in HG(X)PRT proteins, except one motif that is located on a flexible loop distal to the active site. This flexible loop encompasses a SerTyr dipeptide that is found in all HG(X)PRTs for which sequence is available. The recently determined crystal structures of the HGXPRT from Toxoplasma gondii in the absence of ligand and in the presence of product have demonstrated that this flexible loop in the apoenzyme is shifted toward the active

show abstract

Section: Methodsmentioning

confidence: 83%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Effects Of Protein Modifyingmentioning

confidence: 99%

See 2 more Smart Citations

The Conserved Serine-Tyrosine Dipeptide in Leishmania donovani Hypoxanthine-guanine Phosphoribosyltransferase Is Essential for Catalytic Activity

Jardim

Ullman

1997

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…The product from the PCR reaction was double digested with NdeI (from New England Biolabs) and SulI (from BRL) and ligated directly into the 3.2-kb NdeIl MI-digested gel-purified pBAce plasmid (Craig et al, 1991). This construct, referred to as pBAcprt, was first transformed into E. coli strain DH5a (from BRL), and subsequently subcloned into E. coli strain S0606 (Agpt-pro-lac, thi, hpt) provided by Per Nygaard (University Institute of Biological Chemistry B, Copenhagen) (Jochimsen et al, 1975). Expression of the cDNA was induced by the previous procedure (Yuan et al, 1990).…”

Section: Construction Of An Expression Plasmid For Human Hgprtasementioning

confidence: 99%

Differential inhibitory effects of GMP‐2′,3′‐dialdehyde on human and schistosomal hypoxanthine–guanine phosphoribosyltransferases

Kanaaneh

Craig

Wang

1994

European Journal of Biochemistry

View full text Add to dashboard Cite

The hypoxanthine–guanine phosphoribosyltransferase (HGPRTase) of human and the parasitic trematode, Schistosoma mansoni, were expressed at high levels in transformed Escherichia coli in their native forms. Guanosine 2′,3′‐dialdehyde 5′‐phosphate (ox‐GMP) was shown to bind irreversibly to both enzymes in a time‐dependent manner. This binding was stabilized by sodium borohydride reduction, suggesting that a Schiff's base is formed between the dialdehyde groups of ox‐GMP and the amino group of a lysine residue in the enzymes. This linkage formation applies also to inosine 2′,3′‐dialdehyde 5′‐phosphate but not to adenosine 2′,3′‐dialdehyde 5′‐phosphate. GMP was found to be protective against ox‐GMP inactivation and [3H]ox‐GMP labeling of both HGPRTases. 5‐Phosphoribosyl‐1‐diphosphate (PRibPP) also protects human HGPRTase against the ox‐GMP inactivation and [3H]ox‐GMP labeling but provides virtually no protection against the ox‐GMP inactivation and labeling of the schistosomal enzyme, even though PRibPP binds to the latter with a threefold higher affinity. These results imply that PRibPP and ox‐GMP compete with each other for binding to the human HGPRTase but not for binding to the schistosomal enzyme. This discrepancy could be exploited for the purpose of designing selective inhibitors of the schistosomal HGPRTase. Guanosine 2′,3′‐dialdehyde (ox‐guanosine) is nearly as active as ox‐GMP in inhibiting schistosomal HGPRTase but much less potent in inhibiting human HGPRTase, suggesting that ox‐guanosine and ox‐GMP may bind equally well to the parasite enzyme. PRibPP can protect human but not schistosomal HGPRTase against the inactivation by ox‐guanosine. Therefore, ox‐GMP and ox‐guanosine must be forming Schiff's bases with the same amino acid residues in each of the two HGPRTases.

show abstract

Microbial Models and Regulatory Elements in the Control of Purine Metabolism

Gots

Benson

Jochimsen

et al. 1977

Novartis Foundation Symposia

View full text Add to dashboard Cite

Location on the chromosome of Escherichia coli of genes governing purine metabolism

Cited by 119 publications

References 24 publications

The Conserved Serine-Tyrosine Dipeptide in Leishmania donovani Hypoxanthine-guanine Phosphoribosyltransferase Is Essential for Catalytic Activity

The Conserved Serine-Tyrosine Dipeptide in Leishmania donovani Hypoxanthine-guanine Phosphoribosyltransferase Is Essential for Catalytic Activity

Differential inhibitory effects of GMP‐2′,3′‐dialdehyde on human and schistosomal hypoxanthine–guanine phosphoribosyltransferases

Microbial Models and Regulatory Elements in the Control of Purine Metabolism

Contact Info

Product

Resources

About